Cytological Studies in the Genus Trigonella Linn. Kusum Agarwal1 and P. K. Gupta. Department of Agricultural Botany, Meerut University, Meerut, India

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1 Cytologia 48: , 1983 Cytological Studies in the Genus Trigonella Linn. Kusum Agarwal1 and P. K. Gupta Department of Agricultural Botany, Meerut University, Meerut, India Received January 8, 1982 Genus Trigonella L. belongs to tribe Trifolieae of family Fabaceae. It is re presented by about 100 species (Willis 1966), which are mainly distributed in eastern part of Mediterranean region, spreading from north to central Europe and in North Africa and temperate Asia. Though this is an economically important genus, not much cytological studies have been undertaken so far. Only 29% of the species have been reported either for chromosome counts or for karyotype analysis (Chekhov 1932, Fryer 1930, Senn 1938, Singh and Roy 1970, Fernandes and Santos 1971, Singh and Singh 1976). The genus was also subjected for inducing mutations (Raghuvanshi and Singh 1974a, 1974b) and for making interspecific crosses (Singh 1973). In view of this an attempt has been made by us to study meiosis and mitosis, specially with a view to study intraspecific variability. The present paper deals with the results of meiosis and root tip mitosis (including karyotypes) involving seven species of Trigonella L. Material and methods Material was obtained in the form of seeds from various sources (Table 1). Plants were raised in pots at Meerut University Experimental Farm, Meerut. Dor mancy of seeds was broken by rubbing them with glass fibre paper. Voucher spe cimens were prepared and deposited in the Department of Agricultural Botany, Meerut University, Meerut. For meiosis, flower buds were collected in the forenoon, fixed for at least 24 hrs in Carnoy's fluid (absolute alcohol: chloroform: acetic acid, 6: 3: 1) and stored in 70% ethyl alcohol. The anthers were squashed in 2.0% acetocarmine. For mitosis, young and healthy root tips were pretreated with saturated solution of ƒ -bromonaphthalene for about one to two hours, fixed in acetic alcohol (1:3) for 24 hours and squashed in 2.0% acetocarmine. Photomicrographs were taken from temporary preparations and measure ments were made with the help of Olympus micrometer eye piece. TCl % and TF % were calculated as earlier done in Crotalaria (Gupta and Gupta 1978). Results 1. Meiosis Meiosis was studied in six species (Figs. 1-3). Chromosome associatoins and 1 Present address: Department of Botany, R. G. (Post-graduate) College, Meerut, India.

2 Table 1. Sources of material, gametic chromosome numbers and chiasmata frequencies in the genus Trigonella

3 1983 Cytological Studies in the Genus Trigonella Linn. 773 Figs Meiosis in different Trigonella species. 1, T. coerulea (1761), metaphase I, , T. cretica, telophase II. 3, T. foenum-graecum, metaphase I, 8II Mitotic metaphase plates in different Trigonella species. 4, T. calliceras. 5, T. coerulea (1769). 6, T. coerulea (1762). 7, T. coerulea (1763). 8, T. corniculata, 9. T. cretica, 10. T. foenum-graecum, 11. T. hamosa.

4 774 Kusum Agarwal and P. K. Gupta Cytologia 48 chiasmata frequencies are presented in Table 1. All species were diploids having n=8, except T. polycerata where the chromosome number was n=22. Meiosis was completely regular in all the species studied except in T. cretica where dyads, triads and polyads were observed at telophase II (Fig. 2). 2. Mitosis and karyotype Mitosis was studied from nine collections belonging to six species, namely T. calliceras, T. coerulea, T. corniculata, T. cretica, T. foenum-graecum and T. hamosa. Out of these, four collections belonged to T. coerulea. All the species were diploids having 2n=16 (Figs. 4-11). Data on chromosome measurements and other features are presented in Tables 2-4. Idiograms are shown in Figs The chromosomes were designated in the decreasing order of length as 1-8. Depending upon their absolute length, chromosomes were classified into four categories, namely A=more than 4.0ƒÊ; B=3.0ƒÊ-4.0ƒÊ; C=2.0ƒÊ-3.0ƒÊ and D= less than 2.0ƒÊ. The chromosomes were further subdivided according to the position of centro mere and the chromosome formulae are given in Table 4. Discussion A survey of cytological literature published in the genus Trigonella revealed that only 29 of the 100 species known in the genus have been cytologically examined so far. Of these 29 species, 25 are known to have 2n=16, 3 species are known to have 2n=32 and 5 species are known to have 2n=44. This also suggests the presence of chromosomal races in certain species. During the present study, in the genus Trigonella seven species were examined, of which only T. polycerata had 2n=44 (Table 1), the remaining six species having 2n=16. All these reports in the present study thus confirmed the available reports. An important study in the genus Trigonella was earlier conducted by Singh and Roy (1970) and by Singh and Singh (1976). On the basis of interspecific hybridi zation, they believed that chromosomes of T. hamosa, T. corniculata and T. cretica are homologous to each other. A hypothesis of homology is based on pairing be haviour at meiosis in F1 hybrids and differences in karyotypes known between them will have no relevance. Such close homology is known to occur between Lolium species which differ greatly in chromosome lengths and DNA contents, the differences being attributed to repetitive or inert DNA (Gupta and Rees 1975). The pachytene studies of F1 hybrids may be rewarding in this respect (Gupta 1976). Singh (1973) also found interchange differences between species. These interchanges may lead to differences in karyotype asymmetry without altering the total chromosome length or the DNA content. During the present study, six species were examined (Table 2-4) for mitosis and all were found to be diploids having 2n=16. The chromatin length among the Figs Idiograms prepared from mitotic metaphase in different Trigonella species. 12, T. calliceras. 13, T. coerulea (1769). 14, T. coerulea (1762). 15, T. coerulea (1964). 16, T. coerulea (1763). 17, T. corniculata. 18, T. cretica. 19, T. foenum-graecum. 20, T. hamosa.

5 1983 Cytological Studies in the Genus Trigonella Linn. 775

6 776 Kusum Agarwal and P. K. Gupta Cytologia 48 Table 2. Analysis of mitotic chromosomes in different species of Trigonella L. (In each species, the first row represents absolute length of chromosome in ƒê, the second row gives arm ratios; the third row gives relative chromosome length and the fourth row gives TCI% six species examined during the present study was found to be highest in T. foenum graecum (33.05ƒÊ) and lowest in T. cretica (16.36ƒÊ). T. foenum-graecum also had relatively more asymmetric karyotype suggesting that it was an advanced species (Table 5), and that it has evolved due to increase in chromatin content and change in karyotype from symmetric to asymmetric. Such a conclusion receives support

7 1983 Cytological Studies in the Genus Trigonella Linn. 777 from earlier study conducted by Singh and Singh (1976). Another important feature of the present study is the intraspecific variability in T. coerulea for which four accessions ranged from 17.19ƒÊ to 29.98ƒÊ. It is remarkable that such a variability should be found within the same species. To Table 3. Chromosome data in 9 collections of Trigonella L. Table 4. Karyotypic formulae of 9 collections belonging to 6 species of Trigonella (A, B, C, and D represent length of the chromosome in decreasing order; sc used as subscript represents secondary constriction in long arm; superscripts 'm', 'sm', and 'st' represent respectively the median, submedian and subterminal position of centromeres what extent this variability can be attributed to personal error in measurements will be difficult to evaluate, but in view of the same technique used, degree of chromosome condensation being same and the measurements taken with Olympus micrometer eye piece, the degree of error can not be of a high order. Therefore, the intraspecific variability in chromatin length is real rather than apparent and calls for attention

8 778 Kusum Agarwal and P. K. Gupta Cytologia 48 and explanation. It is not surprising, however, in view of the fact that a species is dynamic, always undergoing a process of change. Acknowledgements The authors are grateful to various agencies listed in Table I for the supply of seed material, which made it possible for us to conduct this study. Table 5. Placement of different Trigonella species in 12 categories proposed by Stebbins (1971), on the basis of karyotypes Summary 1. In the genus Trigonella, male meiosis was studied in six species and mitosis with the help of root tip was studied in nine collections belonging to six species. All the species were diploid having n=8 and 2n=16, except T. polycerata where the chromosome number was n= Meiosis was normal in all the cases. Chiasmata frequency/bivalent varied from 1.77 (T. foenum-graecum) to 1.98 (T. corniculata). 3. Total chromatin length varied from 16.36ƒÊ (T. cretica) to 33.05ƒÊ (T. foenum graecum). 4. The intraspecific variability in chromatin length was observed in four accessions of T. coerulea, suggesting that the species was dynamic and always undergoing a process of change. 5. Species were classified according to classification of Stebbins (1971). Only four of the 12 catagories were represented, indicating that the degree of asym metry was low. References Chekhov, V. B Karyosystematical analysis of the tribe Trifolieae DC. Bull. Appl. Bot. Genet. and Pl. Breed. 1: Fernandes, A. and Santos, M. F Contribution a la connaissance cytotaxinomique des spermatophyta de Portugal IV. Leguminosae. Bot. Soc. Brot. 45:

9 1983 Cytological Studies in the Genus Trigonella Linn. 779 Fryer, J. R Cytological studies in Medicago, Melilotus and Trigonella. Can. J. Res. 3: Gupta, P. K Structural basis of quantitative difference in nuclear DNA between Lolium temulantum and L. rigidum. Ind. J. Exp. Biol. 14: and Rees, H Tolerance of Lolium hybrids to quantitative variation in nuclear DNA. Nature 257: 587. Gupta, R. and Gupta, P. K Karyotypic studies in the genus Crotalaria Linn. Cytologia 42: Raghuvanshi, S. S. and Singh, A. K. 1974a. A possible mutagenic effect of colchicine in Tri gonella foenum-graecum. Cytologia 39: and b. Studies on the effect of Gamma-rays on Trigonella foenum-graecum. Cytologia 39: Senn, H. A Chromosome number relationships in the Leguminosae. Bibl. Genet. 12: Singh, A Studies on the interspecific hybrids of Trigonella corniculata L., T. hamosa L. and T. cretica L. Genetica 44: and Roy, R. P Karyological studies in Trigonella, Indigofera and Phaseolus. Nucleus 13: and Singh, D Karyotype studies in Trigonella. Nucleus 19: Stebbins, G. L Chromosomal Evolution in Higher Plants. Edward Arnold (publ.) Ltd., London. Willis, J. C A Dictionary of the Flowering Plants and Ferns, VIII Edition. Cambridge University Press.